Remodeling (turnover) of bone is the process by which the adult skeleton is continually being resorbed (removed) and formed (replaced). Bone remodeling involves the synthesis of bone matrix by osteoblasts and its resorption by osteoclast cells. Both cell types are controlled by osteocytes that express and/or secrete M-CSF, RANKL, sclerostin, and osteoprotegerin regulating bone formation and bone loss (Hughes et al., 2010; Nakashima et al., 2011; Xiong et al., 2011). Increased numbers and/or over-activation of and other defects in osteoclasts can lead to the development of diseases characterized by a net decrease in bone mass or bone malformations. This can eventually result in low bone mass (osteopenia) or osteoporosis. The most common of such diseases, and perhaps the best known, is osteoporosis occurring particularly in women after the onset of menopause. In fact, osteoporosis is the most significant underlying cause of skeletal fractures in late middle-aged and elderly women. While estrogen deficiency has been strongly implicated as a factor in postmenopausal osteoporosis, there is longstanding evidence that remodeling is a locally controlled process that takes place in discrete packets throughout the skeleton. Further diseases associated with bone loss include Paget's disease, periodontal disease, osteosarcoma cancer, bone metastasis and inflammation-associated bone loss like in the context of rheumatoid arthritis, psoriatic arthritis, spondyloarthritis, SLE and systemic sclerosis.
Patients with a decreased bone mass are at a higher risk of invalidity due to fractures and given the rapidly aging population, medical costs have become a serious problem. Currently used therapeutics specifically targeting bone loss, e.g., in the osteoporosis space include bisphosphonates, calcitonin, SERMs, and anti-RANKL antibodies. The gold standard therapy bisphosphonates is taken up by osteoclasts and induces apoptosis; however, one of the pitfalls of biphosphonates include contraindications in case of kidney failure, a common problem in the aging population (e.g., target population of osteoporosis treatment) and the extremely long uptake in bone, which has inherent safety risks. Calcitonin, a natural ligand for its receptor on osteoclasts, inhibits osteoclast function but is, however, second-line choice in patients with long-term corticosteroid therapy. Biological agents like anti-RANKL antibodies are preferred for their high target specificity (WO2012/163887; WO2008/142164).
The development of anti-osteoclastogenic therapeutics is often based on the existing molecules described above by linking them or producing them as small molecules. Other upcoming therapeutics are either chemical-based, natural product-based or biological-based, which render them the qualities of greater cost-effectiveness, safety and target specificity, respectively (Kim et al., 2013). While most of them inhibit osteoclast formation from osteoclast precursors (OCP) to mononuclear osteoclasts (mOC), hence, further influencing fusion and function, therapeutics specifically inhibiting the fusion into multinucleated osteoclasts and, in particular, targeting the fusogenic molecules, are lacking.
Dendritic cell-specific transmembrane protein (DC-STAMP) is essential for cell-cell fusion in osteoclasts (Yagi et al., 2005) and in vitro addition of an antibody blocks formation of multinucleated osteoclasts (WO2010/127180). Although the mechanism is poorly understood, DC-STAMP and its putative ligand might induce some fusogenic molecules as DC-STAMP is shown to have signaling properties. As its expression in dendritic cells (DC) influences differentiation by suppressing granulocyte development, but also phagocytosis to reduce antigen-presenting capability of DCs (Zhang et al., 2014), one must be aware of possible side effects by blocking DC-STAMP.
Desialylation was also shown to block the fusion from TRAP-positive mononuclear cells into multinuclear osteoclasts indicating the essential role of sialic acids. Both alpha (2,6)-linked sialic acid and alpha (2,3)-linked sialic acid are demonstrated on osteoclast precursors and during osteoclast differentiation, but only the alpha (2,6)-linked sialic acid was suggested to have a role in fusion (Takahata et al., 2007).
Although alpha (2,6)-linked sialic acid is a possible ligand for siglec-15, the same group does not particularly put this binding forward in the explanation for the osteoclastogenic properties of siglec-15 (Kameda et al., 2013). They and others (Hiruma et al., 2011; Ishida-Kitagawa et al., 2012; Hiruma et al., 2013) describe the association of siglec-15 with DNAX-activating protein 12 kDa (DAP12), making use of its immunoreceptor tyrosine-based activation motif (ITAM-motif) to co-stimulate the RANK-RANKL pathway. Disruption of this stimulation via siglec-15 in vitro or ex vivo through deficiency or blockage interferes with the development of functional multinucleated osteoclasts (WO2012/045481; M. Stuible et al., 2014). Siglec-15-deficient mice were also shown to be mildly osteopetrotic.
Xiao et al., 2012, explored osteoporosis gene expression profiles in monocytes using a high-throughput microarray platform by grouping individual differentially expressed genes into gene sets and gene ontology terms. A group of 49 genes were assigned in nine clusters using a graph clustering method, including siglec-1 in cluster 1, of which the genes were enriched in similar pathways of immune responses and circulatory system processes. However, Takahata et al., 2007, have appointed a possible sialoadhesin ligand alpha (2,3)-linked sialic acid to only have a minor contribution in fusion of osteoclasts and moreover observed a decreased expression of sialoadhesin with osteoclast differentiation. This led to the assumption that siglec-15, a siglec with a different expression pattern and strikingly different structure and signaling capacity than siglec-1, is the only siglec that is upregulated during osteoclastogenesis (Hiruma et al., 2011; Kameda et al., 2013).